Abstract Hemoglobinopathies, such as sickle cell disease (SCD) and thalassemia, are among the greatest public health concerns in the world. Although new therapeutic modalities, such as gene therapy, are currently being tested, there is a pressing need for pharmacologic approaches to treat general patient populations. Our long-term goal is to develop a compound(s) that induces fetal-type globin (HbF) production by targeting the transcriptional complex regulating globin switching. The objective of this application is to determine molecular mechanisms underlying the LRF-NuRD-mediated ?-globin silencing and identify a mean(s) to target them. Our central hypothesis is that the LRF-containing NuRD complex is a potential target for HbF reactivation therapy. The rationale for the proposed research is that understanding the LRF/NuRD-mediated globin regulation will provide greater understanding of the transcriptional complex regulating ?-globin repression and facilitate development of novel therapeutic strategies for HbF induction therapy. Guided by strong preliminary data, we expect to achieve our objective by pursuing the three specific aims: 1) to determine the molecular basis for ?- globin reactivation in the absence of the LRF/NuRD; 2) to identify a domain(s) of CHD3/4 necessary for ?- globin silencing; and 3) to determine functional significance of LRF/CHD3 interaction in controlling ?-globin silencing. In Aim1, we will employ ChIP-seq and ATAC-seq foot-printing to determine how the NuRD- associated pathways silences ?-globin expression in adult erythroid cells and how ?-globin is induced upon LRF depletion. In Aim2, we will identify a minimal domain(s) of CHD3/4 responsible for ?-globin silencing. To do so, we will perform a functional domain mapping using CRISPR-Cas9 gene mutagenesis. In Aim3, we will determine functional significance of the LRF/CHD3 interaction in controlling ?-globin silencing. Our published work and preliminary data strongly suggest that the NuRD-associated pathways, in which LRF and BCL11A are involved, represent the near entirety of the ?globin-switch?. We expect that the combined approaches proposed here will elucidate the role of LRF and the NuRD complex in ?-globin silencing and facilitate development of novel strategies for HbF reactivation therapy for hemoglobinopathies.